Objective: To characterize the metabolism and uptake of the reduced derivatives of methotrexate, aminopterin and homofolic acid. Approach: Reduced derivatives of methotrexate, aminopterin and homofolic acid, such as the tetrahydro-, 5-methyl- and 5-formyl- tetrahydro-derivatives, with a high specific H3-radio activity (greater than 1 Ci/mmole) will be prepared and chromatographic procedures for the purification and separation of these compounds will be developed. The metabolism and uptake of these compounds, as well as their metabolic action on folate metabolism will be studied in Lactobacillus casei (ATCC 7469) and Pediococcus cerevisiae (ATCC 8081), in L 1210 leukemic cells, resistant and sensitive to methotrexate and in rats. The possible inhibition of mammalian homocysteine 5-methyl-tetrahydrofolic acid methyl transferase by these compounds will be studied. The potential usefullness of these compounds for the chemotherapy of cancer will be discussed in view of the experimental results. The folic acid coenzyme pattern in human tumor tissue and leukemic leukocytes will be determined. Special emphasis will be given to an accurate determination of the intracellular proportion of polyglutamate and 5-methyl-tetrahydro-derivatives of folic acid.